Growth suppression in Salmonella enterica serovar Hadar (S. Hadar) was investigated, in vitro under strict anaerobiosis and in vivo in the intestine of the day-old chicken. Stationary-phase cultures of 20 S. Hadar field strains were tested against each other for growth suppression activity by their ability to suppress the multiplication of low counts of minority cultures inoculated into them as nalidixic acid-resistant mutants. All strains showed profound growth suppression. Four S. Hadar strains were selected and further tested for their ability to suppress growth of S. Enteritidis, S. Typhimurium, S. Virchow and S. Saintpaul. One of the four strains (S. Hadar 18) was randomly selected for further studies. Precolonization of chicken with S. Hadar 18 prevented superinfection with any of the serovars mentioned above. From more than 1000 TnphoA mutants of S. Hadar 18 screened against the parent strain anaerobically in vitro, four were non-suppressive with TnphoA insertions in dapF, aroD, sgaT or tatA. Only the dapF mutant was also non-suppressive in the chicken intestine.